【摘要】 目的了解Survivin启动子在HepG2肝癌细胞与3T3细胞中诱导Trail蛋白表达能力的差异,并评价其驱动Trail诱导肝癌细胞凋亡的能力。方法以pGL3-surp340质粒为模板,扩增Survivin340启动子,构建重组质粒pcDNA3.1/SurP/Trail。将重组质粒分别转染HepG2肝癌细胞和3T3细胞,通过Western Blot检测两种细胞中Trail蛋白表达的差异。利用流式细胞术检测HepG2肝癌细胞及3T3细胞转染重组质粒pcDNA3.1/SurP/Trail后细胞凋亡的情况。结果成功构建了重组质粒pcDNA3.1/SurP/Trail,转染HepG2和3T3两种细胞后,通过Western Blot检测,HepG2肝癌细胞Trail蛋白表达的相对灰度值为0.49±0.43,较3T3细胞Trail蛋白表达(0.15±0.37)明显增强(P<0.05)。利用流式细胞术检测,HepG2肝癌细胞转染重组质粒组的凋亡率(%)为11.59±0.74,空白质粒组为3.45±0.52,未转染组为2.67±0.34;3T3细胞重组质粒组的凋亡率(%)为3.26±0.24,空白质粒组3.16±0.15,未转染组2.83±0.27。HepG2细胞转染重组质粒组凋亡率明显高于其他对照组(P<0.05)。结论含Survivin基因启动子的重组质粒pcDNA3.1/SurP/Trail在肿瘤细胞中能增强Trail蛋白的表达,并能诱导HepG2肝癌细胞凋亡。更多还原

【Abstract】 Objective To investigate the expression difference of Trail protein induced by Survivin promoter in different cells and to evaluate the apoptosis of hepatoma carcinoma cells induced by Trail with Survivin promoter.Methods Using pGL3-surp340 plasmid as template,Survivin340 gene promoter was amplified.The recombinant plasmid pcDNA3.1/SurP/Trail was constructed.The expression differenece of Trail protein between HepG2 cells and 3T3 cells was detected by Western Blotting method.The apoptosis of HepG2 cells and 3T3 cells which transfected with pcDNA3.1/SurP/Trail was analyzed by double labeling flow cytometry.Results The recombinant plasmid pcDNA3.1/SurP/Trail was successfully constructed.Western Blotting results showed that the expression of Trail protein in HepG2 cells(0.49±0.43) was higher than that in 3T3 cells(0.15±0.37),and the apoptosis rate of HepG2 cells transfected with recombinant plasmid(11.59±0.74) was also significantly higher than other groups(P<0.05).Conclusion The recombinant plasmid pcDNA3.1/SurP/Trail containing Survivin promoter enhanced the expression of Trail in tumor cells and induced cell apoptosis of HepG2 cells.更多还原